For measuring epidermal thickness, paraffin sections were stained with H and E. and images were taken by inverted microscopy (Olympus). Epidermal thickness was measured under microscope.
The immunofluorescent (IF) staining of mice skin biopsies for CD31 was performed to assess the microvessel densities. The 5‐μm thick tissues were sectioned from frozen tissue. To inhibit non‐specific antigen–antibody reactions, tissues were blocked with goat serum (Boster Biological Technology) for 30 min and washed for 3 times with PBS buffer. Primary rabbit anti‐CD31 (Abcam) was added and incubated at 4°C overnight. The next day, recovery temperature at 37°C, slides were washed in PBS for 3 times and incubated with secondary antibody (Goat anti‐rabbit IgG; Zhongshanjinqiao) for 2 h at room temperature. Finally, slices were washed for three times with PBS, incubated with 4′, 6‐diamidino‐2‐phenylindole (DAPI; Solarbio) for 10 min at room temperature. Sections were then imaged using LSCM (Olympus FV1200MPE).
The immunofluorescent (IF) staining of mice skin biopsies for CD31 was performed to assess the microvessel densities. The 5‐μm thick tissues were sectioned from frozen tissue. To inhibit non‐specific antigen–antibody reactions, tissues were blocked with goat serum (Boster Biological Technology) for 30 min and washed for 3 times with PBS buffer. Primary rabbit anti‐CD31 (Abcam) was added and incubated at 4°C overnight. The next day, recovery temperature at 37°C, slides were washed in PBS for 3 times and incubated with secondary antibody (Goat anti‐rabbit IgG; Zhongshanjinqiao) for 2 h at room temperature. Finally, slices were washed for three times with PBS, incubated with 4′, 6‐diamidino‐2‐phenylindole (DAPI; Solarbio) for 10 min at room temperature. Sections were then imaged using LSCM (Olympus FV1200MPE).